Abstract: Background　Cathepsin S （CTSS） is a lysosomal cysteine protease which can be found in many kinds of tumor tissues with high level expression. However，the role of CTSS in pathogenesis and development among multiple cancers remain unclarified. Objective　To investigate the difference in the expression of CTSS in pancarcinoma tissues，and to clarify the correlation between CTSS level and tumor progression and prognosis. Methods　RNA sequencing （RNA-Seq） gene expression profiles and clinical data of 33 types of cancerous tissue and normal tissue were downloaded from TCGA and UCSC databases，CTSS gene expression data in normal human tissues were collected from the HPA database. The relationship between CTSS expression and overall survival time （OS） of tumor patients was visually analyzed using R data packets.The correlation between CTSS and tumor mutation burden （TMB），microsatellite instability （MSI） and immune checkpoint was analyzed. Western blot assay was used to detect the difference of CTSS expression between normal glioma cell line （HA1800） and three glioma cell lines （C6，U87，U251）. The effect of CTSS expression on cell proliferation was verified by CCK-8 assay. One-way ANOVA was used to analyze the differences between groups. Results　The levels of CTSS in colon cancer，lung adenocarcinoma，non-small cell lung cancer，pancreatic cancer，prostate cancer and rectal cancer were lower than those in normal tissues，while the levels of CTSS in ovarian cancer，glioblastoma，renal clear cell carcinoma，renal papillary cell carcinoma，gastric adenocarcinoma，thyroid cancer and endometrial cancer were higher than those in normal tissues，with statistical significance （P<0.05）. Survival analysis results showed that in transitional cell carcinoma of bladder，low-grade glioma，ovarian serous cystic adenocarcinoma，sarcomatoid lung cancer，melanoma and uveal melanoma，there were statistically significant differences in OS between patients with high and low expression of CTSS （P<0.05）. Correlation analysis showed that CTSS level was positively correlated with TMB in low-grade glioma，esophageal cancer and breast cancer （P<0.05）. CTSS levels were negatively correlated with TMB in non-small cell lung cancer，lung adenocarcinoma，hepatocellular carcinoma and head and neck carcinoma （P<0.05）. In testicular carcinoma，melanoma，pheochromocytoma，pancreatic cancer，ovarian cancer，non-small cell lung cancer，lung adenocarcinoma，hepatocellular carcinoma，low-grade glioma，renal papillary cell carcinoma，head and neck carcinoma and diffuse large B-cell lymphoma，CTSS levels were negatively correlated with MSI （P<0.05）. The levels of CTSS in 33 tumor tissues were positively correlated with TNFRSF9，CD86，HAVCR2 and CD200R1 （P<0.05）. Western blot results showed that the CTSS level of HA1800 cells was lower than that of the three malignant glioma cells，and the difference was statistically significant（P<0.05）. The results of CCK-8 showed that the proliferation ability of HA1800，C6，U87 and U251 cells in shRNA interference group was lower than that in control group and empty carrier group after 48 and 72 h culture，and the results were statistically significant （P<0.05）. Conclusion　CTSS is associated with a variety of tumor immune checkpoints，MSI and TMB. High expression of CTSS can promote the proliferation of tumor cells and reduce the survival of patients，and can be used as a biomarker for poor prognosis of various tumors.

Key words: Carcinoma, Cathepsin S, Biomarker, Prognosis, Immune infiltration

摘要： 背景　蛋白酶 S（CTSS）是一种溶酶体半胱氨酸蛋白酶，在多种肿瘤组织中表达水平上调，但CTSS水平与肿瘤进展及预后的关系尚不明确。目的　探讨CTSS在泛癌组织中的表达差异，明确CTSS水平与肿瘤进展及预后的相关性。方法　从TCGA数据库中下载33种癌组织RNA-Seq基因表达谱以及相关临床数据，从HPA数据库收集人类正常组织中的CTSS基因表达数据。利用R数据包可视化分析CTSS表达差异及其肿瘤患者总生存时间（OS）之间的关系，探讨CTSS和肿瘤突变负荷（TMB）、微卫星不稳定性（MSI）以及免疫检查点的相关性；采用Western blot法检测正常胶质细胞株（HA1800）及3种胶质瘤细胞株（C6、U87、U251）CTSS表达差异；采用CCK-8法验证CTSS表达对细胞增殖作用的影响，采用单因素方差分析组间差异。结果　结肠癌、肺腺癌、非小细胞肺癌、胰腺癌、前列腺癌和直肠癌组织中CTSS水平低于正常组织，卵巢癌、胶质母细胞瘤、肾透明细胞癌、肾乳头状细胞癌、胃腺癌、甲状腺癌和子宫内膜癌中CTSS表达水平高于正常组织，差异均具有统计学意义（P<0.05）。生存分析结果显示，在膀胱移行细胞癌、低级别胶质瘤、卵巢浆液囊性腺癌、肉瘤样肺癌、黑色素瘤、葡萄膜黑色素瘤中，CTSS高表达与低表达组患者OS比较，差异均具有统计学意义（P<0.05）。相关性分析结果显示，低级别胶质瘤、食管癌、乳腺癌中CTSS水平与TMB呈正相关（P<0.05）；非小细胞肺癌、肺腺癌、肝细胞癌和头颈癌中CTSS水平与TMB呈负相关（P<0.05）；睾丸癌、黑色素瘤、嗜铬细胞瘤、胰腺癌、卵巢癌、非小细胞肺癌、肺腺癌、肝细胞癌、低级别胶质瘤、肾乳头状细胞癌、头颈癌和弥漫性大B细胞淋巴瘤中，CTSS的水平与MSI呈负相关（P<0.05）；33种肿瘤组织中CTSS水平与TNFRSF9、CD86、HAVCR2、CD200R1均呈正相关（P<0.05）。Western blot结果显示HA1800细胞中CTSS水平低于3种恶性胶质瘤细胞，差异有统计学意义（P<0.05）。CCK-8检测结果显示，HA1800、C6、U87、U251细胞培养48和72 h后，shRNA干扰组细胞的增殖能力低于对照组和空载体组，结果均有统计学意义（P<0.05）。结论　CTSS与多种肿瘤免疫检查点、MSI和TMB相关，CTSS高表达可以促进肿瘤细胞增殖，降低患者生存期，可作为多种肿瘤预后不良的生物标志物。

关键词: 癌, 组织蛋白酶 S, 生物标志物, 预后, 免疫浸润